1. Field of the Invention
The present invention relates to monoclonal antibodies directed specifically against an antigen of a protozoan parasitizing a mollusk. It relates more particularly among these monoclonal antibodies to those which are directed against an antigen of a protozoon belonging to the phylum of Ascetospora as defined by LEVINE N. D. et al.((1980) A newly revised classification of the Protozoa., 27, 37-58). And among these antibodies it relates more especially to those which are directed against an antigen of the protozoon Bonamia ostreae, parasite of the flat oyster species Ostrea edulis L.
2. Brief Review of the Prior Art
It is known that mollusks, particularly sea mollusks, can be parasitized by a protozoon. Several pathogenic protozoa have been identified, particularly among the genera Minchinia, Haplosporidium and Marteilia (GRIZEL H. et al. (1974) Science et P eches, Bull. Inst. P eches Marit., No. 240, 7-30; PERKINS F.O. (1979) Mar. Fish Rev., 41 (1-2), 25-37). Another protozoon, Bonamia ostreae, which is the cause of a parasitosis of flat oyster cultivation beds in France has, since it was brought to light (COMPS M. et al. (1980) C. R. Acad. Sc. Paris, 290, ser. D, 383-384), been the subject of numerous studies focusing both on its structure (PICHOT Y. et al. (1980) Rev. Trav. Inst. P eches Marit., 43 (1), 131-140) and on the epidemiologic situation resulting from its presence (TIGE G. et al. (1981) Science et P eche, Bull. Inst. P eches Marit., No. 315, 13-20).
These parasitoses decimate the mollusk populations. They can lead to the disappearance of species. They have, of course, an impact of considerable economic importance when they affect cultivated mollusks (GRIZEL H. (1983) Cons. Int. ExpLor. Met, Gert.: 9, 30 p), especially when they take the form of an endemic epizootic. This is precisely the case in the parasitosis of the oyster Ostrea edulis L. in which the etiologic agent is Bonamia ostreae, in respect of which it has been experimentally observed that a single individual was able to cause the death of an oyster to which it had been inoculated four months earlier. This parasitosis, having been identified successively in various French oyster beds, is now found in certain oyster-farming basins in several European states, particularly Spain, Ireland, the Netherlands and the United Kingdom, and in the United States of America; it may also be noted that at present a protozoon of the genus Bonamia is suspected of being at the origin of a parasitosis of the flat oyster species Ostrea luteria in New Zealand.
The importance of the economic stakes associated with these parasitoses has led to a search for measures making it possible, if not to eradicate them, at least to check them.
These measures are most often of a prophylactic nature when no breed of the mollusk concerned has been found which is resistant to the parasitosis or when no anti-parasitic treatment has proven efficacious. They then consist essentially in preventing the introduction, into healthy cultivation beds, of animals which are already parasitized. They thus require, for their implementation, the availability of an effective method of diagnosis characterized by its ease of use and its reliability.
No such method of diagnosis has been described. In fact, the only known method consists in carrying out histological examinations on the mollusks. This method is poorly suited for a routine practice because of the means required for its implementation and the necessary recourse to a specialist for interpretation of the results. Moreover, strictly speaking it does not permit quantification of the degree of infestation of the parasitized mollusks. Another important disadvantage is the length of time between the preparation of the samples and the availability of the results.
The Applicant has now found a method which is simple to use and reliable and is based on the use of monoclonal antibodies. Given that the mollusks, faced with a parasitic infestation, are unable to develop a humorally-mediated immune response of the antibody type, it appeared in fact to the applicant that it was necessary to demonstrate the presence of the protozoa themselves. Now, a mono-clonal antibody specifically recognising an antigen of a protozoon constitutes a sure means for demonstrating its presence.
The obtaining of monoclonal antibodies recognising an antigen of a protozoon has already been described within the scope of studies focusing on protozoa of the genus Leishmania which, as parasites of human white corpuscles, are, from a phylogenetic point of view, substantially distant from the protozoa parasitizing the mollusks. But the available publications (particularly the international patent application WO 86/02098) do not allow the particular difficulties to be overcome which, associated with the isolation and the purification of a protozoon from mollusks which it has parasitized, did not exist in the case of the protozoa of the genus Leishmania which it is possible to cultivate and for which there exist pure strains in various collections.